The present study demonstrates that antimicrobial whole-flock treatments, combined with footbathing, can be a useful tool to eliminate both clinical signs of footrot and the causative agent, i.e. virulent D nodosus. This makes antimicrobial treatments applicable for elimination programs based on PCR results, always under consideration of the disadvantages of using critically important antimicrobials. Real-time PCR testing proved to be a more sensitive method than clinical examination, as it also identified clinically healthy carriers of D nodosus.
Ideally, samples would have been collected before the elimination process as well, however this was not an experimental but a field study. It remains unclear, whether the aprB2-positive farms never eliminated benign D nodosus in their flocks, due to healthy carriers which never received any treatments, or whether the farmers introduced purchased carriers without clinical signs to the flock which again spread the agent. The latter can be assumed likely in flock 9, as whole flock antimicrobial treatments eliminate both genotypes, and the biosecurity deficits regarding the new outbreak support this hypothesis.
The risk-based pool-of-10 sampling strategy worked well in our setting, since all animals positive for virulent D nodosus were found in the high-risk group. Pooling and pool size were found to be reliable and appropriate. This supports the application of risk-based pooling for flock screenings, elimination and surveillance programs, as it is cost and labour effective.
One major challenge in footrot elimination is the development of diagnostic tests which can discriminate reliably between target and non-target strains . Regional elimination programs are usually aimed at virulent D nodosus, as large-scale elimination of benign D nodosus is not economically justifiable . In Switzerland, the use of the PCR test was found to be the most economic and sensitive way to eliminate footrot in the planned nationwide control program . Moreover, the PCR test can be applied to other ruminants like goats and cattle or camelids, which is important for targeted disease control . In the current study, it detected symptomless carriers of virulent and benign D nodosus before clinical signs were manifest and proved thereby to be more sensitive than clinical scoring. The PCR results agreed with the clinical appearance of virulent and benign strains, including subclinical carriers, in agreement with previous observations [10, 27, 28]. Virulent D nodosus caused progressive underrunning footrot in flock 9 during the outbreak that followed sampling. When the farmer discovered the reinfection, 20% of the flock was already infected and had to be treated. Two weeks earlier, also the farm of origin of the infected ram had noticed a new footrot outbreak.
In flock 24, no clinical signs were observed when the flock was revisited 3 weeks later. It must be noted that the single positive animal in that flock was again the ram. It had a low load of virulent D nodosus based on the high Ct-value and underwent immediate antimicrobial treatment, once the positive result was known.
Likewise, there were no clinical signs in the flocks positive for benign D nodosus at the time of sampling, probably due to dry weather conditions. According to the flock health veterinarian, aprB2-positive D nodosus occasionally caused either mild inflammations (likely in lambs due to increased vulnerability of their interdigital space in rough conditions) or severe inflammations with acute lameness, but no underrunning (H. Strobel, personal communication). It can be concluded that clinical features are consistent with the PCR classification if benign is defined as non-progressive in the South German sheep population. This is in line with findings from Locher et al.  in a previous study on clinically free sheep flocks in Switzerland, was described in Australia by Best et al.  and also reported from Norwegian farms by Vatn et al. . Further, Best et al.  proofed, that real-time PCR was significantly more sensitive in detection of aprV2 in clinically healthy sheep compared with culture/gelatinase test. McPherson et al. stated, that the results of the real-time PCR did not agree sufficiently with the clinical findings in Australian sheep flocks and therefore depended on additional culturing . In Sweden benign D nodosus is often associated with underrunning (score ≥ 3 lesions) . Similarly, clinical impact caused by benign D nodosus was reported from a wild-range alpine ibex colony in Switzerland . Therefore, further research is required to determine whether the PCR classification used in this study corresponds to clinical presentation, if considered as elimination tool in other species like goats or other populations, since the genetic background of the host also plays a role . If clinical disease expression and PCR results match each other, the test is a useful diagnostic tool for early diagnosis of virulent D nodosus at the flock level and can support veterinarians to distinguish between footrot and interdigital inflammations of other cause at an early stage of disease, when clinical signs are not definite . This will lead to reduced antimicrobial use, as non-progressive interdigital inflammations respond well to alternative treatments.
Animal welfare is an important concern in today’s society, and footrot is one of the main welfare issues in sheep production . Antimicrobial whole-flock treatments have shown successful results for elimination of the disease . Strobel et al.  reported cure rates of more than 99% after one or two systemic administrations. Therefore, if a single antimicrobial treatment can improve animal welfare significantly, it should be considered . On the other hand, the World Health Organization considers the popularly used macrolides as “critically important antimicrobials” for human medicine, and gives them highest priority within this category, suggesting to restrict their use if alternative treatments are available . This puts pressure on veterinarians to limit antimicrobial use . Before implementing a footrot elimination program based on antimicrobials, it must be investigated, whether strict biosecurity measures can be fulfilled after the elimination to avoid reinfections, repeated antimicrobial treatments and economic losses. Especially the farmer’s engagement and will to eliminate the disease is mentioned as a key factor for successful elimination [1, 29, 38], besides culling of non-responders and strict quarantine of purchased animals [15, 38]. Our study shows that monitoring the whole flock remains essential even after clinical elimination, and the aprV2/aprB2 PCR is a very useful tool for this purpose.
Whilst macrolide treatments are at present an effective tool to eliminate virulent D nodosus, it is likely, that they will no longer be licensed or effective in the future. Repeated footbathing in 10% ZnSO4 solution proved to be effective for the elimination of virulent strains of D nodosus in a proof-of-concept study . However, since the application of disinfectants based on zinc, copper or formol solution is debatable, further investigation on alternative foot bath solutions and alternative ways to treat clinical footrot is desirable. The reintroduction of virulent or benign D nodosus to a sanitized flock through a clinically healthy carrier must be considered the main risk , and was also described in a field study by Forbes et al. . Rams were found to be a major risk factor as they often change hands and are being shown in sheep markets. The establishment of a footrot free flock certification, which is based on a negative PCR outcome for D nodosus, as Stamphøj et al.  suggested, would allow farmers to select rams from footrot free breeders, which might be a key factor for footrot elimination in European sheep production. Ram breeders would be obliged to join in an accreditation scheme.